Information on EC 3.4.22.32 - Stem bromelain

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The expected taxonomic range for this enzyme is: Ananas comosus

EC NUMBER
COMMENTARY hide
3.4.22.32
-
RECOMMENDED NAME
GeneOntology No.
Stem bromelain
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
broad specificity for cleavage of proteins, but strong preference for Z-Arg-Arg-/-NHMec amongst small molecule substrates
show the reaction diagram
; endopeptidase; The most abundant of the cysteine endopeptidases of the stem of the pineapple plant, Ananas comosus. Distinct from the bromelain found in the pineapple fruit (3.4.22.33). Scarcely inhibited by chicken cystatin and also very slowly inactivated by 3.4.22.4.
-
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hydrolysis of peptide bond
CAS REGISTRY NUMBER
COMMENTARY hide
37189-34-7
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ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
azocasein + H2O
fragments of azocasein
show the reaction diagram
-
-
-
-
Azocoll + H2O
Hydrolyzed azocoll
show the reaction diagram
-
-
-
-
Benzoyl-Arg-Arg 4-nitrophenyl ester + H2O
Benzoyl-Arg-Arg + 4-nitrophenol
show the reaction diagram
-
-
-
-
-
Benzoyl-citrulline 4-nitroanilide + H2O
Benzoyl-citrulline + 4-nitroaniline
show the reaction diagram
-
-
-
-
-
Benzoyl-DL-Ala ethyl ester + H2O
Benzoyl-DL-Ala + ethanol
show the reaction diagram
-
-
-
-
-
Benzoyl-DL-Arg 4-nitroanilide + H2O
Benzoyl-DL-Arg + 4-nitroaniline
show the reaction diagram
-
-
-
-
-
Benzoyl-Gly ethyl ester + H2O
Benzoyl-Gly + ethanol
show the reaction diagram
-
-
-
-
-
Benzoyl-L-Arg amide + H2O
Benzoyl-L-Arg + NH3
show the reaction diagram
-
-
-
-
-
Benzoyl-L-Arg ethyl ester + H2O
Benzoyl-L-Arg + ethanol
show the reaction diagram
-
-
-
-
-
Benzoyl-L-Arg methyl ester + H2O
Benzoyl-L-Arg + methanol
show the reaction diagram
-
-
-
-
-
Benzoyl-Phe-Val-Arg 4-methylcoumarin 7-amide + H2O
?
show the reaction diagram
-
-
-
-
-
benzyl-Phe-Val-Arg-4-nitroanilide + H2O
?
show the reaction diagram
-
-
-
-
?
Benzyloxycarbonyl-Arg-Arg 4-methylcoumarin 7-amide + H2O
Benzyloxycarbonyl-Arg-Arg + 7-amino-4-methylcoumarin
show the reaction diagram
Benzyloxycarbonyl-Arg-Arg 4-nitrophenyl ester + H2O
Benzyloxycarbonyl-Arg-Arg + 4-nitrophenol
show the reaction diagram
-
-
-
-
-
benzyloxycarbonyl-Arg-Arg-NH-4-methylcoumarin 7-amide + H2O
?
show the reaction diagram
-
synthetic substrate
-
?
benzyloxycarbonyl-Arg-Arg-p-nitroanilide + H2O
benzyloxycarbonyl-Arg-Arg + p-nitroanilide
show the reaction diagram
-
-
-
-
?
Benzyloxycarbonyl-citrulline 4-nitroanilide + H2O
Benzyloxycarbonyl-citrulline + 4-nitroaniline
show the reaction diagram
-
-
-
-
-
Benzyloxycarbonyl-Gly 4-nitrophenyl ester + H2O
Benzyloxycarbonyl-Gly + 4-nitrophenol
show the reaction diagram
-
-
-
-
-
Benzyloxycarbonyl-Gly-citrulline 4-nitroanilide + H2O
?
show the reaction diagram
-
-
-
-
-
Benzyloxycarbonyl-Gly-Phe-citrulline 4-methylcoumarin 7-amide + H2O
?
show the reaction diagram
-
-
-
-
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Benzyloxycarbonyl-Gly-Phe-citrulline 4-nitroanilide + H2O
?
show the reaction diagram
-
-
-
-
-
Benzyloxycarbonyl-Gly-Phe-Phe-citrulline 4-nitroanilide + H2O
?
show the reaction diagram
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best substrate
-
-
-
Benzyloxycarbonyl-Gly-Pro-citrulline 4-nitroanilide + H2O
?
show the reaction diagram
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-
-
-
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Benzyloxycarbonyl-L-Ala 4-nitrophenyl ester + H2O
Benzyloxycarbonyl-L-Ala + 4-nitrophenol
show the reaction diagram
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-
-
-
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Benzyloxycarbonyl-L-Asn 4-nitrophenyl ester + H2O
Benzyloxycarbonyl-L-Asn + 4-nitrophenol
show the reaction diagram
-
-
-
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Benzyloxycarbonyl-L-Lys 4-nitrophenyl ester + H2O
Benzyloxycarbonyl-L-Lys + 4-nitrophenol
show the reaction diagram
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-
-
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benzyloxycarbonyl-L-Tyr 4-nitrophenyl ester + H2O
benzyloxycarbonyl-L-Tyr + 4-nitrophenol
show the reaction diagram
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-
-
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Benzyloxycarbonyl-Phe-Arg 4-methylcoumarin 7-amide + H2O
Benzyloxycarbonyl-Phe-Arg + 7-amino-4-methylcoumarin
show the reaction diagram
benzyloxycarbonyl-Phe-Arg-NH-4-methylcoumarin 7-amide + H2O
?
show the reaction diagram
-
synthetic substrate
-
?
Benzyloxycarbonyl-Phe-citrulline 4-nitroanilide + H2O
?
show the reaction diagram
-
-
-
-
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benzyloxycarbonyl-Phe-Val-Arg-NH-4-methylcoumarin 7-amide + H2O
?
show the reaction diagram
-
synthetic substrate
-
?
Benzyloxycarbonyl-Pro-citrulline 4-nitroanilide + H2O
?
show the reaction diagram
-
-
-
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Bz-Phe-Val-Arg-4-nitroanilide + H2O
?
show the reaction diagram
-
-
-
-
?
Bz-Phe-Val-Arg-4-nitroanilide + H2O
Bz-Phe-Val-Arg + 4-nitroaniline
show the reaction diagram
-
-
-
-
?
casein + H2O
?
show the reaction diagram
casein + H2O
hydrolyzed casein
show the reaction diagram
casein + H2O
L-Tyr + ?
show the reaction diagram
-
-
-
-
?
CD25 + H2O
?
show the reaction diagram
Fibrin + H2O
Hydrolyzed fibrin
show the reaction diagram
-
-
-
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Gelatin + H2O
?
show the reaction diagram
-
-
-
-
?
Gly ethyl ester + H2O
Gly + ethanol
show the reaction diagram
-
-
-
-
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Hemoglobin + H2O
?
show the reaction diagram
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-
-
-
?
Hemoglobin + H2O
Hydrolyzed hemoglobin
show the reaction diagram
hemoglobin + H2O
L-Tyr + ?
show the reaction diagram
-
-
-
-
?
Hide powder + H2O
Hydrolyzed hide powder
show the reaction diagram
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-
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Isolated soybean protein + H2O
?
show the reaction diagram
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-
-
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L-Phe ethyl ester + H2O
L-Phe + ethanol
show the reaction diagram
-
-
-
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L-Pyroglutamyl-L-Phe-L-Leu 4-nitroanilide + H2O
?
show the reaction diagram
-
-
-
-
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L-Tyr ethyl ester + H2O
L-Tyr + ethanol
show the reaction diagram
-
-
-
-
-
Meuchenia sp. insoluble muscle protein + H2O
hydrolyzed Meuchenia sp. insoluble muscle protein
show the reaction diagram
-
-
-
-
?
N-alpha-benzyloxycarbonyl-Lys-p-nitrophenyl ester + H2O
N-alpha-benzyloxycarbonyl-Lys + p-nitrophenol
show the reaction diagram
-
-
-
-
?
N-Benzyloxycarbonyl-L-Phe-L-Ser methyl ester + H2O
?
show the reaction diagram
-
-
-
-
-
Nalpha-CBZ-L-lysine 4-nitrophenyl ester + H2O
Nalpha-CBZ-L-lysine + 4-nitrophenol
show the reaction diagram
-
-
-
-
?
P-selectin glycoprotein ligand-1 + H2O
?
show the reaction diagram
-
-
-
-
?
protein + H2O
peptides
show the reaction diagram
Pyr-Phe-Lys-4-nitroanilide + H2O
Pyr-Phe-Lys + 4-nitroaniline
show the reaction diagram
-
isozyme SBA/b is about 3fold less active than isozyme SBA/a
-
?
tert-butyloxycarbonyl-Leu-Arg-Arg-4-methylcoumaryl-7-amide + H2O
tert-butyloxycarbonyl-Leu-Arg-Arg + 7-amino-4-methylcoumarin
show the reaction diagram
-
-
-
?
Tosyl-citrulline 4-nitroanilide + H2O
Tosyl-citrulline + 4-nitroaniline
show the reaction diagram
-
-
-
-
-
Tosyl-L-Arg methyl ester + H2O
Tosyl-L-Arg + methanol
show the reaction diagram
-
-
-
-
-
Tosyl-L-Lys methyl ester + H2O
Tosyl-L-Lys + methanol
show the reaction diagram
-
-
-
-
-
Wheat gluten + H2O
Hydrolyzed wheat gluten
show the reaction diagram
-
-
-
-
-
Z-Arg-Arg-4-nitroanilide + H2O
?
show the reaction diagram
-
-
-
-
?
Z-Arg-Arg-7-amido-4-methylcoumarin + H2O
?
show the reaction diagram
-
-
-
-
?
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
CD25 + H2O
?
show the reaction diagram
-
bromelain proteolytically cleaved cell-surface CD25 from activated CD4+ T cells, a mechanism of action to exert therapeutic benefits in inflammatory conditions, overview
-
-
?
P-selectin glycoprotein ligand-1 + H2O
?
show the reaction diagram
-
-
-
-
?
protein + H2O
peptides
show the reaction diagram
additional information
?
-
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2-hydroxyethyl disulfide
-
-
alpha2-Macroglobulin
-
-
-
Ananas comosus bromelain inhibitor
Cystatin
-
chicken, weak inhibition
-
ethanol
Guanidine-HCl
-
complete loss of activity at 6 M guanidine-HCl
HgCl2
-
-
Human plasma
-
-
-
Inhibitor from pineapple stem
iodoacetic acid
-
-
L-3-carboxy-2,3-trans-epoxypropionylleucylamido(4-guanidino)butane
-
irreversible
phenylmercuric acetate
-
-
Potato cysteine proteinase inhibitor
-
-
-
Rabbit anti-stem bromelain antibodies
-
-
-
Sucrose
-
bromelain in the presence of 1 M sucrose is destabilized under thermal stress, the average melting temperature decrease by 5C, additionally the enzyme is inactivated faster at 60C
Sulfhydryl reagent
-
-
-
sulfur dioxide
-
strong inhibitor
trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane
-
i.e. E-64, slow inactivation
trehalose
-
bromelain in the presence of 1 M trehalose is destabilized under thermal stress, the average melting temperature decrease by 7C, additionally the enzyme is inactivated faster at 60C
additional information
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
2,3-dimercapto-1-propanol
-
not as effective in activation as cysteine
cysteine
-
inactive unless treated with
DTT
-
not as effective in activation as cysteine
Guanidine-HCl
-
at 2 mM
Thioglycol
-
not as effective in activation as cysteine
Urea
-
at 5 M and 8-9 M
additional information
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
210
Benzoyl-Gly ethyl ester
-
-
1.2 - 67
Benzoyl-L-Arg amide
57 - 200
benzoyl-L-Arg ethyl ester
32
benzoyl-L-Arg methyl ester
-
-
0.0097
benzoyl-Phe-Val-Arg 4-methylcoumarin 7-amide
-
-
0.249
benzyl-Phe-Val-Arg-4-nitroanilide
-
tartaric buffer containing 12% ethanol, pH 3.2, 25C, stem bromelain
-
0.0154
benzyloxycarbonyl-Arg-Arg 4-methylcoumarin 7-amide
-
-
0.0074 - 0.0081
benzyloxycarbonyl-Arg-Arg-NH-4-methylcoumarin 7-amide
0.0153
Benzyloxycarbonyl-Gly-Phe-citrulline 4-methylcoumarin 7-amide
-
-
0.0831
Benzyloxycarbonyl-Phe-Arg 4-methylcoumarin 7-amide
-
-
0.0554 - 0.0636
benzyloxycarbonyl-Phe-Arg-NH-4-methylcoumarin 7-amide
0.003 - 0.0039
benzyloxycarbonyl-Phe-Val-Arg-NH-4-methylcoumarin 7-amide
0.03919 - 0.305
Bz-Phe-Val-Arg-4-nitroanilide
0.129 - 5.26
casein
0.4 - 2.42
L-Pyroglutamyl-L-Phe-L-Leu 4-nitroanilide
0.53
N-Benzyloxycarbonyl-L-Phe-L-Ser methyl ester
-
-
84
tosyl-L-Lys ethyl ester
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-
additional information
additional information
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.55
acetyl-Gly ethyl ester
Ananas comosus
-
-
0.21 - 0.36
Benzoyl-Gly ethyl ester
0.0035 - 0.027
Benzoyl-L-Arg amide
0.39 - 0.5
benzoyl-L-Arg ethyl ester
0.11
benzoyl-L-Arg methyl ester
Ananas comosus
-
-
0.893
benzoyl-Phe-Val-Arg 4-methylcoumarin 7-amide
Ananas comosus
-
-
20
benzyl-Phe-Val-Arg-4-nitroanilide
Ananas comosus
-
tartaric buffer containing 12% ethanol, pH 3.2, 25C, stem bromelain
-
27
benzyloxycarbonyl-Arg-Arg 4-methylcoumarin 7-amide
Ananas comosus
-
-
0.024 - 0.04
benzyloxycarbonyl-Arg-Arg-NH-4-methylcoumarin 7-amide
1.75 - 3.35
Benzyloxycarbonyl-Gly 4-nitrophenyl ester
0.12
Benzyloxycarbonyl-Gly-Phe-citrulline 4-methylcoumarin 7-amide
Ananas comosus
-
-
2.5 - 8.3
benzyloxycarbonyl-L-Ala 4-nitrophenyl ester
1.4 - 1.5
Benzyloxycarbonyl-L-Asn 4-nitrophenyl ester
Ananas comosus
-
-
7.4 - 9.8
benzyloxycarbonyl-L-Lys 4-nitrophenyl ester
0.4 - 0.44
benzyloxycarbonyl-L-Tyr 4-nitrophenyl ester
0.13
Benzyloxycarbonyl-Phe-Arg 4-methylcoumarin 7-amide
Ananas comosus
-
-
0.49 - 6.08
benzyloxycarbonyl-Phe-Arg-NH-4-methylcoumarin 7-amide
5.1 - 6.4
benzyloxycarbonyl-Phe-Val-Arg-NH-4-methylcoumarin 7-amide
0.29 - 21.45
Bz-Phe-Val-Arg-4-nitroanilide
0.41 - 18.08
casein
0.68 - 3.94
L-Pyroglutamyl-L-Phe-L-Leu 4-nitroanilide
3.4
N-Benzyloxycarbonyl-L-Phe-L-Ser methyl ester
Ananas comosus
-
-
0.035
Tosyl-L-Lys methyl ester
Ananas comosus
-
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.1
chicken cystatin
-
both isozymes, pH 6.0, 25C
-
0.145
SDS
-
25C
additional information
additional information
-
k2/Ki for E-64
-
IC50 VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.55
SDS
Ananas comosus
-
25C
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.17
-
partially purified enzyme, 37C, pH 6.5
0.62
-
purified enzyme, substrate casein
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
3.2
-
assay at
4 - 4.5
-
L-pyroglutamyl-L-Phe-L-Leu 4 nitroanilide, enzyme form F4 and F5
5 - 8
-
wide plateau, Nalpha-benzoyl-L-Arg amide
5
-
denatured hemoglobin, enzyme form F4
6 - 7
-
substrate-dependent
7 - 8.5
-
casein
8
-
native enzyme, at 35C
10
-
randomly polymer-coupled enzyme preparations, at 35C
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5 - 10
-
5: about 40% of activity maximum, 10.0: about 70% of activity maximum
5 - 11
-
pH 5.0: native enzyme shows about 25% of maximal activity, enzyme covalently coupled to the CNBr-activated Sepharose shows about 30% of maximal activity, enzyme affinity-bound to a Sepharose matrix precoupled with the lactin concanavalin A shows about 35% of maximal activity, pH 11: native enzyme shows about 25% of maximal activity, enzyme covalently coupled to the CNBr-activated Sepharose shows about 30% of maximal activity, enzyme affinity-bound to a Sepharose matrix precoupled with the lactin concanavalin A shows about 60% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
60 - 70
-
randomly polymer-coupled enzyme preparations
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4.8
-
isoelectric focusing, isozyme SBA/b, and isozyme SBA/a variant 2
4.9
-
isoelectric focusing, isozyme SBA/a variant 1
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
18000
-
Ananas comosus, enzyme form I-2, II-1 and II-2, gel filtration
18800
-
Ananas comosus, bromelain A, gel filtration
19700
-
Ananas comosus, bromelain B, gel filtration
22500
-
Ananas comosus, gel filtration
23430
-
Ananas comosus, enzyme form F9, mass spectroscopy
23550
-
isozyme SBA7A, mass spectroscopy
23560
-
isozyme SBA/b, mass spectroscopy
23800
-
SDS-PAGE
24400
-
Ananas comosus, enzyme form F4, mass spectroscopy
24470
-
Ananas comosus, enzyme form F5, mass spectroscopy
26000
-
SDS-PAGE
28400
-
Ananas comosus, sedimentation velocity and equilibrium ultracentrifugation experiments
33000
-
Ananas comosus, determination from sedimentation and diffusion coefficient
35300
-
Ananas comosus, sedimentation velocity measurements
37000
-
Ananas comosus, gel filtration
43000
-
SDS-PAGE, recombinant protein
additional information
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
monomer
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.8 - 2
-
stem bromelain at pH 2.0 is maximally unfolded and characterized by significant loss of secondary structure (about 80%) and almost complete loss of tertiary contacts, at pH 0.8 a molten globule state is observed with secondary structure content similar to that of native protein but no tertiary structure
680545
2
-
almost complete loss of native tertiary contacts, acid unfolded state
664339
3 - 9
-
4C, 24 h, stable
30285
3.2 - 7
-
stem bromelain solubilized at pH 7.0 and at pH 3.2 retains, after 16 h at 256C, 10 and 40% of the initial activity, respectively
731526
4 - 10
-
5C, 24 h, stable
30273
5
-
50C, 60 min, stable
30285
5.5 - 10
-
stem bromelain is fully resistant against urea around neutral pH (5.5 to 10.0) and unfolds only below pH 5.0
707537
5.6 - 9
-
stable
30297
7 - 10
-
from pH 7.0 to 10.0, the protein's secondary structure remains the same, although a slight loss of tertiary structure is observed. Above pH 10.0, there is a significant and irreversible loss of secondary and tertiary structure. At pH 10.0, SBM shows a significant increase in 8-anilino-1-naphthalene-sulfonate binding relative to the native state. No significant loss of activity is observed up to pH 10.0, beyond which there is an irreversible loss of activity
707283
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4
-
recombinant protein stored at 4C shows a loss in enzymatic activity. The percentage loss in recombinant bromelain enzymatic activity is 27.5%
40
-
unstable above, crude enzyme extract
50
-
pH 5, 60 min, stable
55
-
pH 6.1, 20 min, 50% loss of activity
63
-
thermal unfolding of native SBM occurs at 63C
63.4
-
melting temperature of the native enzyme
70
soluble enzyme retains 53% activity after incubation at 70C, immobilized enzyme retains over 70% activity after incubation at 70C
100
-
recombinant bromelain shows no activity after 30 min of incubation at 100C
additional information
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
bromelain immobilized to IDA-Sepharose 6B matrix loaded with Cu2+, Ni2+ and Zn2+ is more resistant to thermal inactivation, as evidenced by retention of over 50% activity after incubation at 60C
-
Casein stabilizes against heat inactivation
-
denaturation in guanidine hydrochloride, the deglycosylated enzyme is more sensitive than the glycosylated one, midpoints of transition are at 2.28 M and 2.86 M, respectively
-
enzymatic activity of bromelain remains uninfluenced by the immobilization of heparin on it
-
enzyme affinity-bound to a Sepharose matrix precoupled with the lactin concanavalin A is more stable to thermal inactivation than native enzyme or enzyme covalently coupled to the CNBr-activated Sepharose
-
glutaraldehyde-crosslinked bromelain has comparable activity to the native enzyme, is more stable against urea, guanidine hydrochloride and temperature-induced inactivation and exhibit better storage ability compared to the unmodified protease
-
glycerol and sorbitol are acting as stabilizers at all concentrations while sucrose and trehalose are destabilizers at lower concentrations, however, act as stabilizers at higher concentrations. Urea and guanidine hydrochloride are denaturants except at lower concentrations
-
immobilization on amino-Sepharose leads to higher proteolytic activity and remarkably enhanced thermal stability as compared to soluble bromelain and that coupled to CNBr- activated Sepharose
immobilized bromelain shows a higher half-life respect to the free enzyme. The addition of free cysteine during immobilization phase, improves bromelain half-life more than 4folds
-
increase in ionic strength by addition of salts results in folded structures somewhat different from the native enzyme. Salt-induced intermediates are characterized by increase in helical content and a significantly reduced exposure of hydrophobic clusters relative to the state at pH 2.0. Salt-induced state shows non-cooperative thermal denaturation alcohol-induced intermediates of the enzyme exhibit increased helical content. Alcohol-induced state shows a cooperative thermal transition
-
Lyophilization causes 27% loss of activity
-
poly(ethylene glycol)-400-induced state has characteristics of molten globule, higher molecular weight poly(ethylene glycol)s cause unfolding of the acid unfolded state
-
Stem bromelain covalently coupled to a thermosensitive polymer of N-isopropylacrylamide either through the amino groups of the enzyme (randomly coupled) or via the lone oligosaccharide chain (uniformly coupled) shows better thermostability. The enzyme coupled via the oligosaccharide chain exhibits better access to the substrate casein as compared to the preparation in which the amino groups formed the point of contact between the enzyme and the polymer.
-
the glycosylated enzyme is more stable than the deglycosylated one
-
the low bromelain activity, particularly in the upper portion of the murine gastrointestinal tract, following oral administration, suggests that extensive gastric inactivation of bromelain may occur in vivo. When formulated in antacid, oral bromelain retains substantial proteolytic activity throughout the gastrointestinal tract (of mice)
-
the pyromellitic anhydride- and poly(maleic anhydride)-modified bromelain does not lose its activity significantly
-
treatment with 10-30% (v/v) 1,1,1,3,3,3-hexafluoroisopropanol induces the partially folded intermediate to adopt much of the native proteins secondary structure, but only a rudimentary tertiary structure, characteristic of the molten globule state. Addition of slightly higher concentrations of 1,1,1,3,3,3-hexafluoroisopropanol causes transformation from an alpha-helix to a beta-sheet and induces formation of a compact nonnative structure
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ORGANIC SOLVENT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
2,2,2-trifluoroethanol
-
the enhanced binding of 1-anilino-8-naphthalene sulfonic acid to the specific/pre-molten globule (SMG) state of stem bromelain upon addition of 30% 2,2,2-trifluoroethanol suggests the presence of a large number of solvent-accessible non-polar clusters in the treated SMG. The formation of an molten globe (MG)-like state characterized by disordered side chain interactions but with considerable secondary structure when the specific/pre-molten globule (SMG) state of stem bromelain is subjected to 30% 2,2,2-trifluoroethanol (TFE). The TFE-induced MG conformation at alkaline pH could represent the conformation that allows stem bromelain to traverse membranes
Ethanol
guanidine-HCl
Methanol
-
25% v/v, 25C, 20 min, stable
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
oxidation of the active site thiol leads to the corresponding sulfinic acid, which is catalytically inactive, irreversible oxidative degradation may lead to reduced activity of isozyme SBA/b with substrate Pyr-Phe-Lys-4-nitroanilide
-
653021
photosensitized oxidation of stem bromelain in the presence of methylene blue results in partial loss of the enzymatic activity even if the essential sulfhydryl group is protected against oxidation
-
30274, 30286
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20C, stored dry, stable
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
2 enzyme forms with pI of 9.45 (basic form) and pI 4.7 (acidic form)
-
2 forms: SB1 and SB2
-
2 isozymes SBA/a and SBA/b
-
6 enzyme forms: SBB 1-5 and SBA
-
affinity chromatography
-
bromelain A and B
-
bromelain from pineapple stem and skin are recovered by a PEG 4000/phosphate aqueous two-phase systems (ATPs) liquid-liquid extraction
-
carboxymethylcellulose 52 column chromatography, and Sephadex G-100 gel filtration
-
CM-52 cellulose column chromatography
-
enzyme forms: I-1, I-2, II-1, II-2
-
native enzyme from stem, reversible inactivation of the protease in 10 mM Na-phosphate, pH 7.5 containing 5 mM of Na-tetrathionate, during purification, reactivation by 8 mM of DTT and 4 mM of EDTA in the same buffer at 25C, followed by gel filtration
-
Ni2+-coated IMAC Hypercel column chromatography
-
to homogeneity
-
using Ni-NTA chromatography
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
cloned into the pENTR/TEV/D-TOPO vector, then sub-cloned into the pDEST17 expression vector. Expression in Escherichia coli BL21
-
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
agriculture
-
a concentration of 0.0003 mM of bromelain is sufficient for 90% growth inhibition of Fusarium verticillioides. Bromelain also inhibits the growth of Fusarium oxysporum f. sp. melonis and Fuarium proliferatum. The enzyme shows a potential use as an effective agent for crop protection
food industry
medicine